There is an urgent need for the development of new anti-HIV drugs against targets different than those currently applied in HAART namely HlV reverse transcriptase and HIV protease. Because of the high mutability of HIV, because of complications with compliance with drug administration and because of complications from drug side effects. virernia due to HIV escape mutants does appear in a significant fraction of patients on HAART therapy. HIV relies on numerous cellular enzymes for replication and proliferation. In principle, the development of drugs against cellular targets would potentially eliminate concern about viral escape mutants and the problems with resistant strains of HIV-1 and HIV-2 in patients, infected in other parts of the world. Candidate cellular targets that show promise include the co-receptors for HIV infectivity and peptides that block interaction of the HIV envelope glycoproteins with CD4 receptor. We have been researching a different candidate cellular targets - the cellular protease required for the cleavage of HIV-gp160 envelope precursor protein to its gp120 and gp41 subunits. In the absence of this intracellular cleavage event virions with uncleaved gpl60 or reduced density of cleaved envelope glycoproteins are unable to infect their target cells. We identified the cellular protease. PC6 to be responsible for gp160 cleavage and ha\e shown that depletion or deletion of this enzyme is sufficient for eliminating HIV infectivity, thereby blocking AIDS disease progression and proliferation of the virus. However, studies demonstrating the role of PC6 on HIV infectivity have been performed exclusively on isolated cells (including primary human T cells and T cell lines) grown in culture. The risks of targeting drugs to deplete or reduce the activity of the cellular PC6 protease are addressed by the studies in this proposal. Deletion of this cellular protease does not affect cell viability. PC6 knockout mice will he generated to assess whether this protease is essential to normal mouse development, to immune system function and to normal homeostasis. We will also identify normal cellular precursor substrates for PC6 activity in human T cells by the powerful proteomics approach. Since PC6 activity does not need to be completely eliminated in order to disable HIV infectivity (due to the requirement for collaboration by eighteen gp120-gp41 subunits for HIV infection of the target cells), then the identification of cellular substrates will serve as a therapeutic index as to how specific and how effective the PC6 protease inhibitors work in isolated cells and in patients. Therefore, this work will provide valuable weight to the argument for targeting drugs to inhibit the cellular PC6 protease, which will inbe helpfulthe development of these inhibitor drugs as an alternative, or complement to existing HAART therapies.